The current projects represent the continuing development and expansion of our research program into the pathophysiology of vaso-occlusion and builds upon the historical strengths of our Center. With this proposal, we now move forward into two novel directions that explore new questions in sickle cell disease and strengthen collaborations between existing projects. Dr. Kohn will address an entirely new research objective for this Center in vitro and in vivo models of human erythropoiesis. Dr. Donald Kohn, one of the leading pediatric gene therapy investigators in this country, has developed a new method to expand stem cells from patients with sickle cell disease into mature erythrocytes or granulocytes in quantities sufficient to allow Dr. Meiselman to make biorheologic measurements of these cells prior to exposure to the in vivo environment and to allow Dr. Coates to study the intrinsic properties of sickle neutrophils. This approach will not only allow Dr. Meiselman to determine if b-globin related differences in sickle RBC rheologic properties are present in virginal sickle RBC, but also to measure the direct rheologic consequences of the genetic manipulations of the RBC proposed by Dr. Kohn. The novel model of human sickle erythropoiesis in the bnx-mouse will provide an in vivo environment for future exploration of hypothesis posed in other Projects. From the collaborations between Drs. Coates, Kalra and Meiselman, we have strong evidence that sickle erythrocytes specifically activate leukocytes which can then participate in endothelial damage. Dr. Coates Project will address the role that the reticuloendothelial systems, a major target in this disease, plays in disease manifestations. Two projects explore the intrinsic properties of the sickle RBC whole two other projects explore those characteristics of the vascular endothelium and reticuloendothelial system which contribute to vaso-occlusion. This closely knit ensemble of project directors have extensive histories of prior NIH support and bring expertise from wide-ranging backgrounds in basic research to bear on the problem of vaso-occlusion.